You had thought about a flow cytometric technique?

Thank you, but I am not sure that FC will work  here. First, as far as I remember male cells differ from female cells only by 2% of DNA. I do  not think that it is enough to separate  them effectively. Second, we have to test some very early stages of development, before egg laying, when the embryo counts only tens or a few hundred cells, and it would be problematic to load them into the FC for separation.   

Garner at 2006 reviewed the sexing process by FC in mammalians, maybe it can serve to you, Regards. 

Unfortunately, it does not work in birds.

Please check out our paper on female specific ubiquitous hintw expression:

https://www.researchgate.net/publication/260559242_HINTW_a_W-chromosome_histidine_triad_nucleotide_binding_protein_%28HINT%29_gene_in_chick_is_expressed_ubiquitously_and_is_a_robust_female_cell_marker_applicable_in_intra-specific_chimera_studies

we know that zga in G. gallus starts at late EGK-II/early EGK-III (a few dozen cells). So your theory can be tested by looking at embryos of late EGK-III (about 200 cells) to EGK-V (about 2000 cells) stages, using the hintw probe.

Article HINTW, a W-Chromosome HINT Gene in Chick, Is Expressed Ubiqu...

Thank you very much Dr. Guojun Sheng. Your method  is very timely and I hope on your technical support. Fortunately,  I came across your publication just after my question was published. And once again if you have any doubts  or questions about the hypothesis you are welcome, we can discuss them online. Not everything was published in the paper.  

I couldn't find HINTW in zebra finch library. How do you think will the primers presented in your paper,  work for ZF also? Thank you.